Nitrogen Fixation
All living organisms require nitrogen. it is a main component of amino acids, nucleic acids (DNA and RNA) and many other important molecules in living cells. However, the majority of organisms cannot utilize nitrogen in the atmosphere directly, and can only utilize nitrogen compounds of certain forms. The processes of forming such nitrogen compounds are collectively referred to as “nitrogen fixation”. In nitrogen fixation, nitrogen is combined with other elements and is “fixed” within nitrogen containing compounds.
Agricultural plants are grown in large quantities all over the world, and substantial amount of nitrogen in soil is consumed by these plants every year. If there were no replenishment of nitrogen, its content in the soil would decrease and this would affect the output of the agricultural plants.
In reality, the soil acquires replenishment of nitrogen element via two routes: the application of nitrogen-containing fertilizers, and the biological nitrogen fixation. It was estimated in the 1980s that the nitrogen fertilizers applied every year globally contain around 8×107 tons of nitrogen, while as much as 4×108 tons of nitrogen is provided by the nature via biological nitrogen fixation in the mean time.
Thus, if nitrogen fixation mechanism can be applied in food crops like wheat and rice plant to allow them to fix nitrogen on their own, the global requirement of nitrogen fertilizers would be greatly reduced and the agricultural output would be increased. This would be very meaningful in solving global food issues and in protecting the eco-environment.
Nitrogen Fixing Microorganisms
Biological nitrogen fixation is mainly accomplished by nitrogen fixing microorganisms.
Nitrogen fixing microorganism can be divided into two major classes: symbiotic and free-living. Typical symbiotic. nitrogen fixing microorganisms include rhizobia that fix nitrogen only when they are in symbiotic relationship with legumes. Free-living nitrogen fixing microorganisms are soilborne microorganisms that can fix nitrogen independently, mainly bacteria and Cyanobacterium (also known as blue-green algae). Common nitrogen fixing bacteria include aerobic Pasteurella, anaerobic Klebsiella, as well as Rhodospirillum and Chromatium that are capable of photosynthesis. Currently, the most common free-living nitrogen fixing bacterium that is used as a nitrogen fertilizer is the aerobic Azotobacter chroococcum. 
Nitrogen Fixation Genes
Among various nitrogen-fixing microorganisms, the most extensively studied one is Klebsiella pneumoniae. Its nitrogen fixation genes comprise 17-20 nif genes, in sequence are J, H, D, K, T, Y, E, N, X, U, S, V, W. Z, M, F, L, A, B, and Q. These genes form 7 operons, listed below (Qi Cheng, Perspectives in Biological Nitrogen Fixation Research. Journal of Integrative Plant Biology, 2008):
NifJ operon: comprises nifJ gene
NifHDKY operon: comprises nifH, nifD, nifK, and nifY gene
NifENX operon: comprises nifE, nifN, and nifX gene
NifUSVM: operon: comprises nifU, nifS, nifV, and nifM gene
NifF operon: comprises nifF gene
NifLA operon: comprises nifL and nifA gene
NifBQ operon: comprises nifB and nifQ gene
The nitrogen fixation system is conserved among all nitrogen fixing microorganisms and the nitrogen fixation genes share very high homology. For example, the nif gene of the nitrogen fixation system of rhizobia is homologous to the nif gene of K. pneumoniae. 
The Heterogeneous Expression of Nitrogen Fixation Genes of Klebsiella pneumoniae 
A 24 kb nif nitrogen fixation gene (NCBI Accession number X13303) is cut off from the chromosome of Klebsiella pneumoniae, and is ligated as a whole into a vector for transforming E. coil (Escherichia coil) to confer the host cell the ability of fixing nitrogen (Ray Dixon, Frank Cannon, Construction of a P plasmid carrying nitrogen fixation genes from Klebsiella pneumoniae. Nature, 1976). However, a yeast transformed with the same gene produce only some but not all of the proteins of the nitrogen fixing enzymes, and is not able to fix nitrogen (Ada Zamir, Stable chromosomal integration of the entire nitrogen fixation acme cluster from Klebsiella pneumoniae in yeast Proc. NatL Acad. Sci. USA, 1981). It can then be seen that it is difficult to express prokaryotic nitrogen fixation genes in eukaryotes to allow the biological nitrogen fixation of eukaryotes such as wheat and rice plants.